title: Discovery and annotation of small proteins using genomics, proteomics, and computational approaches.
abstract: Small proteins (10-200 amino acids [aa] in length) encoded by short open reading frames (sORF) play important regulatory roles in various biological processes, including tumor progression, stress response, flowering, and hormone signaling. However, ab initio discovery of small proteins has been relatively overlooked. Recent advances in deep transcriptome sequencing make it possible to efficiently identify sORFs at the genome level. In this study, we obtained ~2.6 million expressed sequence tag (EST) reads from Populus deltoides leaf transcriptome and reconstructed full-length transcripts from the EST sequences. We identified an initial set of 12,852 sORFs encoding proteins of 10-200 aa in length. Three computational approaches were then used to enrich for bona fide protein-coding sORFs from the initial sORF set: (1) coding-potential prediction, (2) evolutionary conservation between P. deltoides and other plant species, and (3) gene family clustering within P. deltoides. As a result, a high-confidence sORF candidate set containing 1469 genes was obtained. Analysis of the protein domains, non-protein-coding RNA motifs, sequence length distribution, and protein mass spectrometry data supported this high-confidence sORF set. In the high-confidence sORF candidate set, known protein domains were identified in 1282 genes (higher-confidence sORF candidate set), out of which 611 genes, designated as highest-confidence candidate sORF set, were supported by proteomics data. Of the 611 highest-confidence candidate sORF genes, 56 were new to the current Populus genome annotation. This study not only demonstrates that there are potential sORF candidates to be annotated in sequenced genomes, but also presents an efficient strategy for discovery of sORFs in species with no genome annotation yet available.
authors.affiliation: Biosciences Division, Oak Ridge National Laboratory, Oak Ridge, TN 37831, USA. yangx@ornl.gov
authors.forename: Xiaohan
authors.initials: X
authors.lastname: Yang
authors.forename: Timothy J
authors.initials: TJ
authors.lastname: Tschaplinski
authors.forename: Gregory B
authors.initials: GB
authors.lastname: Hurst
authors.forename: Sara
authors.initials: S
authors.lastname: Jawdy
authors.forename: Paul E
authors.initials: PE
authors.lastname: Abraham
authors.forename: Patricia K
authors.initials: PK
authors.lastname: Lankford
authors.forename: Rachel M
authors.initials: RM
authors.lastname: Adams
authors.forename: Manesh B
authors.initials: MB
authors.lastname: Shah
authors.forename: Robert L
authors.initials: RL
authors.lastname: Hettich
authors.forename: Erika
authors.initials: E
authors.lastname: Lindquist
authors.forename: Udaya C
authors.initials: UC
authors.lastname: Kalluri
authors.forename: Lee E
authors.initials: LE
authors.lastname: Gunter
authors.forename: Christa
authors.initials: C
authors.lastname: Pennacchio
authors.forename: Gerald A
authors.initials: GA
authors.lastname: Tuskan
mesh_terms: D019295:Computational Biology; D020224:Expressed Sequence Tags; D023281:Genomics; D058977:Molecular Sequence Annotation / Q000379:methods*; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D018515:Plant Leaves / Q000235:genetics; D010940:Plant Proteins / Q000235:genetics; D032107:Populus / Q000235:genetics; D040901:Proteomics; D022661:RNA, Untranslated / Q000235:genetics; D012107:Research Design
chemical_list: D010940:Plant Proteins; D022661:RNA, Untranslated